Little is known how patterns of cross-over (CO) numbers and distribution during meiosis are established. Here, we reveal that cyclin-dependent kinase A;1 (CDKA;1), the homolog of human Cdk1 and Cdk2, is a major regulator of meiotic recombination in Arabidopsis. Arabidopsis plants with reduced CDKA;1 activity experienced a decrease of class I COs, especially lowering recombination rates in centromere-proximal regions. Interestingly, this reduction of type I CO did not affect CO assurance, a mechanism by which each chromosome receives at least one CO, resulting in all chromosomes exhibiting similar genetic lengths in weak loss-of-function cdka;1 mutants. Conversely, an increase of CDKA;1 activity resulted in elevated recombination frequencies. Thus, modulation of CDKA;1 kinase activity affects the number and placement of COs along the chromosome axis in a dose-dependent manner.

The N‐end rule pathway is a highly conserved constituent of the ubiquitin proteasome system, yet little is known about its biological roles.Here we explored the role of the N‐end rule pathway in the plant immune response. We investigated the genetic influences of components of the pathway and known protein substrates on physiological, biochemical and metabolic responses to pathogen infection.We show that the glutamine (Gln) deamidation and cysteine (Cys) oxidation branches are both components of the plant immune system, through the E3 ligase PROTEOLYSIS (PRT)6. In Arabidopsis thaliana Gln‐specific amino‐terminal (Nt)‐amidase (NTAQ1) controls the expression of specific defence‐response genes, activates the synthesis pathway for the phytoalexin camalexin and influences basal resistance to the hemibiotroph pathogen Pseudomonas syringae pv tomato (Pst). The Nt‐Cys ETHYLENE RESPONSE FACTOR VII transcription factor substrates enhance pathogen‐induced stomatal closure. Transgenic barley with reduced HvPRT6 expression showed enhanced resistance to Ps. japonica and Blumeria graminis f. sp. hordei, indicating a conserved role of the pathway.We propose that that separate branches of the N‐end rule pathway act as distinct components of the plant immune response in flowering plants.

Dynamic regulation of protein function and abundance plays an important role in virtually every aspect of plant life. Diversifying mechanisms at the RNA and protein level result in many protein molecules with distinct sequence and modification, termed proteoforms, arising from a single gene. Distinct protein termini define proteoforms arising from translation of alternative transcripts, use of alternative translation initiation sites, and different co- and post-translational modifications of the protein termini. Also site-specific proteolytic processing by endo- and exoproteases generates truncated proteoforms, defined by distinct protease-generated neo-N- and neo-C-termini, that may exhibit altered activity, function, and localization compared with their precursor proteins. In eukaryotes, the N-degron pathway targets cytosolic proteins, exposing destabilizing N-terminal amino acids and/or destabilizing N-terminal modifications for proteasomal degradation. This enables rapid and selective removal not only of unfolded proteins, but also of substrate proteoforms generated by proteolytic processing or changes in N-terminal modifications. Here we summarize current protocols enabling proteome-wide analysis of protein termini, which have provided important new insights into N-terminal modifications and protein stability determinants, protein maturation pathways, and protease–substrate relationships in plants.

In plants, the protein RPM1-INTERACTING PROTEIN4 (RIN4) is a central regulator of both pattern-triggered immunity and effector-triggered immunity. RIN4 is targeted by several effectors, including the Pseudomonas syringae protease effector AvrRpt2. Cleavage of RIN4 by AvrRpt2 generates potentially unstable RIN4 fragments, whose degradation leads to the activation of the resistance protein RESISTANT TO P. SYRINGAE2. Hence, identifying the determinants of RIN4 degradation is key to understanding RESISTANT TO P. SYRINGAE2–mediated effector-triggered immunity, as well as virulence functions of AvrRpt2. In addition to RIN4, AvrRpt2 cleaves host proteins from the nitrate-induced (NOI) domain family. Although cleavage of NOI domain proteins by AvrRpt2 may contribute to pattern-triggered immunity regulation, the (in)stability of these proteolytic fragments and the determinants regulating their stability remain unexamined. Notably, a common feature of RIN4, and of many NOI domain protein fragments generated by AvrRpt2 cleavage, is the exposure of a new N-terminal residue that is destabilizing according to the N-end rule. Using antibodies raised against endogenous RIN4, we show that the destabilization of AvrRpt2-cleaved RIN4 fragments is independent of the N-end rule pathway (recently renamed the N-degron pathway). By contrast, several NOI domain protein fragments are genuine substrates of the N-degron pathway. The discovery of this set of substrates considerably expands the number of known proteins targeted for degradation by this ubiquitin-dependent pathway in plants. These results advance our current understanding of the role of AvrRpt2 in promoting bacterial virulence.

Toxic proteins are prime targets for molecular farming (the generation of pharmacologically active or biotechnologically usable compounds in plants) and are also efficient tools for targeted cell ablation in genetics, developmental biology, and biotechnology. However, achieving conditional activity of cytotoxins and maintaining the toxin-expressing plants as stably transformed lines remain challenging. Here, we produce a switchable version of the highly cytotoxic bacterial RNase barnase by fusing the protein to a portable protein degradation cassette, the low-temperature degron cassette. This method allows conditional genetics based on conditional protein degradation via the N-end rule or N-degron pathway and has been used to vice versa accumulate and/or deplete a diverse variety of highly active, unstable or stable target proteins in different living multicellular organisms and cell systems. Moreover, we expressed the barnase fusion under control of the trichome-specific TRIPTYCHON promoter. This enabled efficient temperature-dependent control of protein accumulation in Arabidopsis (Arabidopsis thaliana) leaf hairs (trichomes). By tuning the levels of the protein, we were able to control the fate of trichomes in vivo. The on-demand formation of trichomes through manipulating the balance between stabilization and destabilization of barnase provides proof of concept for a robust and powerful tool for conditional switchable cell arrest. We present this tool as a potential strategy for the manufacture and accumulation of cytotoxic proteins and toxic high-value products in plants or for conditional genetic cell ablation.